Three-dimensional optically cleared tissue imaging for analyzing endoscopic images of gastrointestinal neoplasms (with video).

Abstract

Objectives: To develop a procedure that matches magnifying endoscopic images with narrow-band imaging to 3D tissue structures using a tissue-clearing technique and to qualitatively and quantitatively analyze specified structures in gastrointestinal neoplasms.

Methods: Endoscopically resected formalin-fixed paraffin-embedded gastrointestinal tissues (three esophagus, four stomach, seven colon) were made transparent by ethyl cinnamate. They were then subjected to fluorescent staining of nuclei and blood vessels followed by 3D imaging using a confocal laser scanning microscope. A one-to-one correspondence between magnifying endoscopic and 3D reconstructed images was established using vessels and crypts with characteristic shapes as guides, and the depth and caliber of specified vessels were measured.

Results: All tissues were optically cleared, which allowed 3D visualization of vascular structures and nuclei in all layers. In the esophagus, intraepithelial papillary capillary loops and subepithelial capillary networks were identified. In the upper part of the stomach, polygonal subepithelial capillary loops surrounding the pits were observed, while in the lower part, surface epithelium with ridge-like structures and coiled vessels were observed. A honeycomb pit structure and surrounding vascular structures were identified in the colon. Quantitative analysis showed the various contrasts of a single continuous vessel in the endoscopic image were due to different depths at which the vessel tortuously ran.

Conclusion: We established a procedure to allow one-to-one correspondence between magnifying endoscopic and 3D reconstructed images and to measure the depth and caliber of endoscopically visualized vessels of interest. This method is expected to improve endoscopic diagnosis and further the development of endoscopic imaging technologies.